Overexpression of multimerin-2 promotes cutaneous melanoma cell invasion and migration and is associated with poor prognosis

doi:10.12122/j.issn.1673-4254.2025.07.14

Abstract

Abstract:

Objective To investigate the inhibitory effect of multimerin-2 (MMRN2) overexpression on growth and metastasis of cutaneous melanoma cells. Methods Clinical data of patients with cutaneous melanoma were obtained from the GEO database to compare MMRN2 expressions between normal and tumor tissues. A protein-protein interaction network was constructed using the STRING database, and the intersecting genes from GEPIA2.0 were subjected to GO and KEGG enrichment analysis. The prognostic relevance of MMRN2 expression level was assessed using Cox regression and "timeROC". The correlations of MMRN2 expression level with immune infiltration and angiogenesis-related genes were analyzed using GSCA database and the ssGSEA algorithm. Colony-forming assay, Transwell assay, and wound healing assay were used to examine the changes in proliferation and migration of cultured cutaneous melanoma cells following MMRN2 knockdown. In a mouse model bearing cutaneous melanoma xenograft, the effect of MMRN2 knockdown on vital organ pathologies, survival of the mice and GM-CSF, CXCL9, and TGF‑β1 protein expressions were analyzed. Results MMRN2 was significantly upregulated in metastatic cutaneous melanoma (P<0.001). Protein interaction network analysis identified 15 intersecting genes, which were enriched in endothelium development and cell-cell junctions. In patients with cutaneous melanoma, a high MMRN2 expression was correlated with a poor prognosis, an advanced T stage, a greater Breslow depth, and ulceration (P<0.05). MMRN2 expression level was strongly correlated with 24 immune cell types (P<0.001), fibroblasts, endothelial cells, and expressions of the pro-angiogenic genes (KCNJ8, SLCO2A1, NRP1, and COL3A1; P<0.001). In cultured B16F10 cells, MMRN2 knockdown significantly suppressed cell proliferation, migration and invasion and caused remo-deling of the immunosuppressive microenvironment. Conclusion MMRN2 overexpression drives progression of cutaneous melanoma by enhancing tumor metastasis, angiogenesis and immune evasion, highlighting its potential as a therapeutic target for melanomas.

Key words: MMRN2, melanoma, prognosis, bioinfo-rmatics, immune infiltration

Jinlong PANG, Xinli ZHAO, Zhen ZHANG, Haojie WANG, Xingqi ZHOU, Yumei YANG, Shanshan LI, Xiaoqiang CHANG, Feng LI, Xian LI. Overexpression of multimerin-2 promotes cutaneous melanoma cell invasion and migration and is associated with poor prognosis[J]. Journal of Southern Medical University, 2025, 45(7): 1479-1489.

Figures/Tables 8

Fig.1 MMRN2 expression in normal and cutaneous melanoma tissues. A: Differential gene expression analysis between tumor and normal tissues using data from GSE46517, GSE83583, and GSE4587 datasets. B: MMRN2 expression levels are significantly higher in metastatic cutaneous melanoma than in normal tissues. *P<0.05, **P<0.01 vs normal tissue; ***P<0.001 vs cutaneous melanoma.

Fig.2 Cutaneous melanoma-related genes and the protein-protein interaction network. A: Intersection analysis of genes associated with cutaneous melanoma using the GEPIA2.0 database. B: Protein-protein interaction (PPI) network constructed by retrieving 30 interacting molecules from the STRING database, resulting in 15 hub genes shared across analyses. C: Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analyses of the hub genes.

Fig.3 Association of MMRN2 expression levels with clinicopathological features of patients with cutaneous melanoma. A: Kaplan-Meier survival analysis of patients with cutaneous melanoma stratified by MMRN2 expression level. B: Correlations of MMRN2 expression level with pathological T stage, Breslow depth and melanoma ulceration in patients with cutaneous melanoma. *P<0.05.

Fig.5 Immune infiltration analysis of MMRN2 in cutaneous melanoma. A: Correlation lollipop chart between MMRN2 and immune infiltration matrix data in TCGA database. The results are visualized using the ggplot2 package. The 24 types of immune cells included aDC (activated DC), B cells, CD8 T cells, cytotoxic cells, DC, eosinophils, iDC (immature DC), macrophages, mast cells, neutrophils, NK CD56bright cells, NK CD56dim cells, NK cells, pDC (plasmacytoid DC), T cells, T helper cells, Tcm (T central memory), Tem (T effector memory), TFH (T follicular helper), Tgd (T gamma delta), Th1 cells, Th17 cells, Th2 cells, and TReg. B: Scatter plot of the correlation between MMRN2 expression level and iDC abundance. C: Scatter plot of correlations between MMRN2 and immune cells in cutaneous melanoma. D, E: Scatter plots of correlations between MMRN2 and tumor fibroblasts in cutaneous melanoma. F: Scatter plot of correlations between MMRN2 and endothelial cells in cutaneous melanoma. Data source: TIMER database, http://timer.comp-genomics.org/timer/

Fig.6 Immune infiltration analysis of MMRN2 in cutaneous melanoma. A: GSVA-based immune cell enrichment scores of MMRN2 in cutaneous melanoma. B: Immune infiltration profiles between high and low MMRN2 expression groups. #: FDR<0.05, *P<0.05, **P<0.01, ***P<0.001 vs low MMRN2 expression group.

Fig.7 Correlation analysis of MMRN2 and angiogenesis-related genes. A: MMRN2 expression is positively correlated with the majority of angiogenesis-associated genes. B: MMRN2 expression level is strongly correlated with KCNJ8, SLCO2A1, NRP1 and COL3A1.

Fig.8 Knockdown of MMRN2 suppresses proliferation, invasion, and migration of cutaneous melanoma cells. A: Western blotting of MMRN2 proteins. B: Colony formation assay for assessing cell proliferation. C: Transwell assay for assessing cell migration and invasion (Original magnification: ×10). D: Knockdown of MMRN2 impairs wound healing in B16 melanoma cells (×200). **P<0.01.

Fig.9 Knockdown of MMRN2 attenuates pathological manifestations and extends survival of mice bearing cutaneous melanoma xenograft. A: HE staining of the liver, spleen, lung, and kidney of mice in MMRN2 knockdown group (×10). B: Kaplan-Meier curve. C: GM-CSF, CXCL9, and TGF-β1 expression levels measured by ELISA. ***P<0.001 vs NC group.

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